Without Its N-Finger, the Main Protease of Severe Acute Respiratory Syndrome Coronavirus Can Form a Novel Dimer through Its C-Terminal Domain

Abstract

ABSTRACT The main protease (M pro ) of severe acute respiratory syndrome coronavirus (SARS-CoV) plays an essential role in the extensive proteolytic processing of the viral polyproteins (pp1a and pp1ab), and it is an important target for anti-SARS drug development. It was found that SARS-CoV M pro exists in solution as an equilibrium of both monomeric and dimeric forms, and the dimeric form is the enzymatically active form. However, the mechanism of SARS-CoV M pro dimerization, especially the roles of its N-terminal seven residues (N-finger) and its unique C-terminal domain in the dimerization, remain unclear. Here we report that the SARS-CoV M pro C-terminal domain alone (residues 187 to 306; M pro -C) is produced in Escherichia coli in both monomeric and dimeric forms, and no exchange could be observed between them at room temperature. The M pro -C dimer has a novel dimerization interface. Meanwhile, the N-finger deletion mutant of SARS-CoV M pro also exists as both a stable monomer and a stable dimer, and the dimer is formed through the same C-terminal-domain interaction as that in the M pro -C dimer. However, no C-terminal domain-mediated dimerization form can be detected for wild-type SARS-CoV M pro . Our study results help to clarify previously published controversial claims about the role of the N-finger in SARS-CoV M pro dimerization. Apparently, without the N-finger, SARS-CoV M pro can no longer retain the active dimer structure; instead, it can form a new type of dimer which is inactive. Therefore, the N-finger of SARS-CoV M pro is not only critical for its dimerization but also essential for the enzyme to form the enzymatically active dimer.